*Target (protein/gene name):pteridine reductase 1 *NCBI Gene # or RefSeq#:LmjF23.0270 *Protein ID (NP or XP #) or Wolbachia#: Q01782 *Organism (including strain):Leishmania major Etiologic Risk Group (see link below): People in areas of North Africa, Middle East and some parts of China. It infects people who are exposed to sand flies or work outside and without fly net. */Disease Information (sort of like the Intro to your Mini Research Write up): Link to TDR Targets page (if present): http://tdrtargets.org/targets/view?gene_id=21977 Link to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.) Essentiality of this protein: Essential in some stages of the organism.
Is it a monomer or multimer as biological unit? (make prediction athttp://www.ebi.ac.uk/msd-srv/prot_int/pistart.html): Predicted to be Multimer Complex of proteins?:Forms a complex of proteins Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism): Yes TDR targets give it a .7 druggability.
*EC#: 1.5.1.33 Link to BRENDA EC# page:http://www.brenda-enzymes.org/php/result_flat.php4?ecno=1.5.1.33 --Show screenshot of BRENDA enzyme mechanism schematic Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):Biopterin + NADPH(need to confirm) -- link to Sigma (or other company) page for assay (see Sigma links below): -- -or link (or citation) to paper that contains assay information -- links to assay reagents (substrates) pages. http://www.sigmaaldrich.com/catalog/product/sigma/b2517?lang=en®ion=US --- List cost and quantity of substrate reagents, supplier, and catalog #:(Sigma $82.30 for 5 MG), maybe Cayman Chemical 10007662 $31 for 5 MG
Structure (PDB or Homology model)
-- PDB # or closest PDB entry if using homology model: 3H4V
-- For Homology Model option:(no need for homology model because have original protein)
---- Show pairwise alignment of your BLASTP search in NCBI against the PDB
---- Query Coverage:
---- Max % Identities:
---- % Positives
---- Chain used for homology:
Current Inhibitors: 25 Expression Information (has it been expressed in bacterial cells): yes http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3904060/ Purification Method: n/a Image of protein (PyMol with features delineated and shown separately): (PDB ribbon image) *Amino Acid Sequence (paste as text only - not as screenshot or as 'code'): MTAPTVPVALVTGAAKRLGRSIAEGLHAEGYAVCLHYHRSAAEANALSATLNARRPNSAI TVQADLSNVATAPVSGADGSAPVTLFTRCAELVAACYTHWGRCDVLVNNASSFYPTPLLR NDEDGHEPCVGDREAMETATADLFGSNAIAPYFLIKAFAHRVAGTPAKHRGTNYSIINMV DAMTNQPLLGYTIYTMAKGALEGLTRSAALELAPLQIRVNGVGPGLSVLVDDMPPAVWEG HRSKVPLYQRDSSAAEVSDVVIFLCSSKAKYITGTCVKVDGGYSLTRA *length of your protein in Amino Acids: 288aa Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 30408.6 Molar Extinction coefficient of your protein at 280 nm wavelength: 27765 TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it. *CDS Gene Sequence (paste as text only): ATGACTGCTCCGACCGTGCCGGTGGCGTTGGTAACAGGCGCCGCGAAGCGTCTTGGCCGC AGTATCGCTGAGGGACTCCACGCGGAGGGGTACGCTGTCTGCTTGCACTATCATCGCTCT GCTGCAGAAGCGAACGCACTATCCGCGACGCTCAACGCAAGGCGACCGAACAGCGCCATC ACGGTGCAGGCGGATCTGAGCAACGTTGCCACAGCCCCGGTCAGCGGCGCTGATGGCTCT GCACCTGTTACCCTCTTCACGCGCTGTGCTGAGTTGGTGGCTGCGTGCTACACCCACTGG GGACGCTGCGACGTGCTAGTGAACAACGCCTCTTCTTTCTACCCCACGCCGCTGCTGAGG AATGACGAGGATGGACACGAGCCCTGTGTCGGAGATAGAGAGGCAATGGAGACGGCCACC GCTGACCTCTTCGGCTCCAACGCGATAGCGCCCTACTTCTTGATTAAGGCGTTCGCGCAT CGCGTCGCGGGCACCCCAGCCAAGCATCGCGGCACCAACTACTCCATCATCAACATGGTC GACGCCATGACGAACCAGCCTCTTCTCGGGTACACCATATATACCATGGCCAAAGGGGCG TTGGAGGGGCTGACACGGTCTGCCGCGCTGGAGCTTGCGCCGCTGCAGATTCGAGTGAAC GGCGTTGGTCCGGGTTTGTCGGTGCTCGTCGATGACATGCCCCCTGCTGTGTGGGAGGGC CACCGCAGCAAGGTGCCTCTGTACCAGCGCGATTCCTCCGCCGCAGAGGTGAGCGACGTT GTTATCTTTCTGTGCTCCTCCAAGGCCAAGTACATCACCGGCACCTGTGTCAAAGTGGAT GGTGGCTACAGCCTTACCCGGGCCTGA *GC% Content for gene: 61.7 % *CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): ATGACTGCGCCGACTGTCCCGGTTGCGCTGGTCACGGGTGCAGCGAAACGTCTCGGTCGT TCTATCGCCGAAGGTCTGCACGCGGAAGGTTACGCAGTTTGCCTGCACTATCACCGTAGC GCAGCGGAAGCGAATGCGCTGTCTGCGACGCTGAATGCACGTCGTCCGAACTCTGCGATC ACGGTACAGGCGGACCTGTCTAACGTTGCGACTGCCCCGGTTTCTGGTGCGGACGGTTCT GCCCCTGTTACCCTGTTCACCCGTTGCGCGGAACTCGTTGCGGCGTGCTACACCCACTGG GGTCGTTGTGACGTCCTGGTTAACAACGCGTCTTCTTTCTACCCGACGCCGCTCCTCCGT AACGACGAAGACGGTCACGAACCTTGCGTTGGCGACCGTGAAGCGATGGAAACCGCGACG GCGGATCTGTTCGGCTCTAACGCAATCGCGCCGTACTTCCTGATCAAAGCGTTCGCCCAC CGTGTTGCGGGTACTCCGGCGAAACACCGTGGTACTAACTACTCTATCATCAACATGGTT GACGCGATGACCAATCAGCCGCTGCTGGGTTATACCATCTACACCATGGCGAAAGGTGCG CTCGAGGGTCTGACTCGTTCTGCGGCGCTGGAACTGGCGCCTCTGCAAATTCGTGTTAAT GGTGTTGGTCCGGGTCTGTCTGTTCTCGTTGACGACATGCCACCGGCCGTTTGGGAAGGC CACCGCTCTAAAGTCCCGCTGTACCAGCGTGACTCTTCTGCTGCAGAAGTTTCCGATGTT GTTATCTTCCTGTGCTCTTCTAAAGCGAAATACATTACGGGTACCTGCGTTAAAGTTGAC GGTGGTTACTCTCTGACCCGTGCGTAA
*GC% Content for gene (codon optimized): 57.5
Do Not Need this info for Spring (but still copy these lines to your Target page for now) Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers): (link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences): Reverse Primer tail
**
TAT CCA CCT TTA CTG TTA CGC ACG GGT CAG AGA GTA AC
*NCBI Gene # or RefSeq#: LmjF23.0270
*Protein ID (NP or XP #) or Wolbachia#: Q01782
*Organism (including strain): Leishmania major
Etiologic Risk Group (see link below): People in areas of North Africa, Middle East and some parts of China. It infects people who are exposed to sand flies or work outside and without fly net.
*/Disease Information (sort of like the Intro to your Mini Research Write up):
Link to TDR Targets page (if present): http://tdrtargets.org/targets/view?gene_id=21977
Link to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.)
Essentiality of this protein: Essential in some stages of the organism.
Is it a monomer or multimer as biological unit? (make prediction at http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html): Predicted to be Multimer
Complex of proteins?:Forms a complex of proteins
Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism): Yes TDR targets give it a .7 druggability.
*EC#: 1.5.1.33
Link to BRENDA EC# page:http://www.brenda-enzymes.org/php/result_flat.php4?ecno=1.5.1.33
-- Show screenshot of BRENDA enzyme mechanism schematic
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents): Biopterin + NADPH(need to confirm)
-- link to Sigma (or other company) page for assay (see Sigma links below):
-- -or link (or citation) to paper that contains assay information
-- links to assay reagents (substrates) pages.
http://www.sigmaaldrich.com/catalog/product/sigma/b2517?lang=en®ion=US
--- List cost and quantity of substrate reagents, supplier, and catalog #: (Sigma $82.30 for 5 MG), maybe Cayman Chemical 10007662 $31 for 5 MG
Structure (PDB or Homology model)
-- PDB # or closest PDB entry if using homology model: 3H4V
-- For Homology Model option:(no need for homology model because have original protein)
---- Show pairwise alignment of your BLASTP search in NCBI against the PDB
---- Query Coverage:
---- Max % Identities:
---- % Positives
---- Chain used for homology:
Current Inhibitors: 25
Expression Information (has it been expressed in bacterial cells): yes
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3904060/
Purification Method: n/a
Image of protein (PyMol with features delineated and shown separately):
(PDB ribbon image)
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
MTAPTVPVALVTGAAKRLGRSIAEGLHAEGYAVCLHYHRSAAEANALSATLNARRPNSAI
TVQADLSNVATAPVSGADGSAPVTLFTRCAELVAACYTHWGRCDVLVNNASSFYPTPLLR
NDEDGHEPCVGDREAMETATADLFGSNAIAPYFLIKAFAHRVAGTPAKHRGTNYSIINMV
DAMTNQPLLGYTIYTMAKGALEGLTRSAALELAPLQIRVNGVGPGLSVLVDDMPPAVWEG
HRSKVPLYQRDSSAAEVSDVVIFLCSSKAKYITGTCVKVDGGYSLTRA
*length of your protein in Amino Acids: 288aa
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 30408.6
Molar Extinction coefficient of your protein at 280 nm wavelength: 27765
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
*CDS Gene Sequence (paste as text only):
ATGACTGCTCCGACCGTGCCGGTGGCGTTGGTAACAGGCGCCGCGAAGCGTCTTGGCCGC AGTATCGCTGAGGGACTCCACGCGGAGGGGTACGCTGTCTGCTTGCACTATCATCGCTCT GCTGCAGAAGCGAACGCACTATCCGCGACGCTCAACGCAAGGCGACCGAACAGCGCCATC ACGGTGCAGGCGGATCTGAGCAACGTTGCCACAGCCCCGGTCAGCGGCGCTGATGGCTCT GCACCTGTTACCCTCTTCACGCGCTGTGCTGAGTTGGTGGCTGCGTGCTACACCCACTGG GGACGCTGCGACGTGCTAGTGAACAACGCCTCTTCTTTCTACCCCACGCCGCTGCTGAGG AATGACGAGGATGGACACGAGCCCTGTGTCGGAGATAGAGAGGCAATGGAGACGGCCACC GCTGACCTCTTCGGCTCCAACGCGATAGCGCCCTACTTCTTGATTAAGGCGTTCGCGCAT CGCGTCGCGGGCACCCCAGCCAAGCATCGCGGCACCAACTACTCCATCATCAACATGGTC GACGCCATGACGAACCAGCCTCTTCTCGGGTACACCATATATACCATGGCCAAAGGGGCG TTGGAGGGGCTGACACGGTCTGCCGCGCTGGAGCTTGCGCCGCTGCAGATTCGAGTGAAC GGCGTTGGTCCGGGTTTGTCGGTGCTCGTCGATGACATGCCCCCTGCTGTGTGGGAGGGC CACCGCAGCAAGGTGCCTCTGTACCAGCGCGATTCCTCCGCCGCAGAGGTGAGCGACGTT GTTATCTTTCTGTGCTCCTCCAAGGCCAAGTACATCACCGGCACCTGTGTCAAAGTGGAT
GGTGGCTACAGCCTTACCCGGGCCTGA
*GC% Content for gene: 61.7 %
*CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only):
ATGACTGCGCCGACTGTCCCGGTTGCGCTGGTCACGGGTGCAGCGAAACGTCTCGGTCGT TCTATCGCCGAAGGTCTGCACGCGGAAGGTTACGCAGTTTGCCTGCACTATCACCGTAGC GCAGCGGAAGCGAATGCGCTGTCTGCGACGCTGAATGCACGTCGTCCGAACTCTGCGATC ACGGTACAGGCGGACCTGTCTAACGTTGCGACTGCCCCGGTTTCTGGTGCGGACGGTTCT GCCCCTGTTACCCTGTTCACCCGTTGCGCGGAACTCGTTGCGGCGTGCTACACCCACTGG GGTCGTTGTGACGTCCTGGTTAACAACGCGTCTTCTTTCTACCCGACGCCGCTCCTCCGT AACGACGAAGACGGTCACGAACCTTGCGTTGGCGACCGTGAAGCGATGGAAACCGCGACG GCGGATCTGTTCGGCTCTAACGCAATCGCGCCGTACTTCCTGATCAAAGCGTTCGCCCAC CGTGTTGCGGGTACTCCGGCGAAACACCGTGGTACTAACTACTCTATCATCAACATGGTT GACGCGATGACCAATCAGCCGCTGCTGGGTTATACCATCTACACCATGGCGAAAGGTGCG CTCGAGGGTCTGACTCGTTCTGCGGCGCTGGAACTGGCGCCTCTGCAAATTCGTGTTAAT GGTGTTGGTCCGGGTCTGTCTGTTCTCGTTGACGACATGCCACCGGCCGTTTGGGAAGGC CACCGCTCTAAAGTCCCGCTGTACCAGCGTGACTCTTCTGCTGCAGAAGTTTCCGATGTT GTTATCTTCCTGTGCTCTTCTAAAGCGAAATACATTACGGGTACCTGCGTTAAAGTTGAC
GGTGGTTACTCTCTGACCCGTGCGTAA
*GC% Content for gene (codon optimized): 57.5
Do Not Need this info for Spring (but still copy these lines to your Target page for now)
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):
Reverse Primer tail
**
TAT CCA CCT TTA CTG TTA CGC ACG GGT CAG AGA GTA AC
Forward primer design
TAC TTC CAA TCC ATG ACT GCG CCG ACT GTC